Diabetes mellitus results from loss or dysfunction of the insulin-producing beta-cells in the pancreas. Despite refined insulin injection regimens, diabetic patients suffer from long-term complications, such as blindness and kidney failure. An ultimate cure for diabetes could be achieved through the generation of replacement insulin-producing cells. To develop these replacement cells, we need to identify the molecular pathways that initiate beta-cell formation and insulin-production. Using genetically engineered mouse models, this proposal examines the role of NKX6 class transcription factors in beta-cell differentiation and function. The specific hypothesis is that different NKX6 transcription factors partially compensate for each other's function in pancreatic cell differentiation. This hypothesis is based on the observation that beta-cell numbers are diminished in Nkx6.1 mutant mice, while Nkx6.1/Nkx6.2 double mutant mice show a reduction in both insulin producing beta- and glucagon-producing alpha-cells. Experiments are proposed to dissect the role of NKX6 factors in pancreatic endocrine development using compound mouse mutants for Nkx6 genes. Aim 1 is to define the role of NKX6 factors in the beta-cell differentiation pathway by attempting to restore beta-cell development in Nkx6.1 mutant mice with different transgenes. Aim 2 examines in mice if pancreatic progenitors are reverted into alternate cellular fates in the absence of NKX6 activity. Aim 3 focuses on the role of NKX6 factors in adult beta-cell function. Using selective inactivation of Nkx6 genes in beta-cells, it will be studied if NKX6 factors control aspects of beta-cell function, such as insulin synthesis or insulin secretion.